Individual Cortical Layers Research Articles

Laminar fMRI in the locked-in stage of amyotrophic lateral sclerosis shows preserved activity in layer Vb of primary motor cortex

Amyotrophic lateral sclerosis (ALS) affects the cerebral cortex layer-dependently, most notably by the foremost targeting of upper motor neurons (UMNs) sited in layer Vb. Previous studies have shown a retained ability of paralysed patients to activate cortical motor networks, even in late-stage ALS. However, it is currently unknown whether such activation reflects a retained capacity to process sensorimotor inputs or if it is a result of actual motor output. Given the distinct function of individual cortical layers, layer-specific functional measurements may provide insight to this question. In this study, using submillimetre resolution laminar fMRI, we assessed the layer-dependent activation associated with attempted (motor) and passive (somatosensory) movements in a locked-in stage ALS patient. We found robust activation in both superficial and deep layers of primary motor cortex (M1). The peak activation in deep layers was localised to layer Vb. These findings demonstrate preserved activity in deep output layers of M1, possibly reflecting a retained ability to engage surviving UMNs despite years of paralysis. Our study underscores the capacity of laminar fMRI to discern subtle cortical activity and elucidates a promising pathway for probing in vivo human ALS pathology with unprecedented resolution.

COMPARATIVE ANALYSIS OF FRACTAL DIMENSIONS OF HUMAN CEREBELLUM: IMPACT OF IMAGE PREPROCESSING AND FRACTAL ANALYSIS METHODS

The aim: To compare the values of the fractal dimensions of human cerebellum obtained using different algorithms of image preprocessing and different methods of fractal analysis. Materials and methods: The study involved 120 people without structural changes in the brain (age 18-86 years, 55 men and 65 women). T1- and T2-weighted MR brain images were studied. Fractal analysis was performed using box counting and pixel dilatation methods. Fractal dimensions of cerebellar tissue as a whole, cerebellar cortex and its individual layers, cerebellar white matter were measured and compared to each other and to fractal dimension of cerebellar white matter determined in cadaveric cerebella. Results: It was no significant difference between fractal dimension values of cerebellar tissue as a whole measured on T1 and T2 weighted magnetic resonance images of cerebellum, and fractal dimension values measured on the same images using different methods of fractal analysis – pixel dilatation and box counting. T2 weighted images are preferable for fractal analysis of different components of cerebellar tissue. Segmentation according to pixel luminance is the preferable image preprocessing method for fractal analysis of cerebellar cortex as a whole, individual cortical layers and cerebellar tissue as a whole; skeletonizing of cerebellar magnetic resonance images is the preferable method of the image preprocessing for fractal analysis of cerebellar white matter. Conclusions: The algorithm of image preprocessing, magnetic resonance imaging sequence and method of fractal analysis should be chosen according to aim of quantitative study of cerebellar magnetic resonance images and features of the studied structure of cerebellum.

Dynamic Contrast Optical Coherence Tomography reveals laminar microvascular hemodynamics in the mouse neocortex in vivo

Studies of flow-metabolism coupling often presume that microvessel architecture is a surrogate for blood flow. To test this assumption, we introduce an in vivo Dynamic Contrast Optical Coherence Tomography (DyC-OCT) method to quantify layer-resolved microvascular blood flow and volume across the full depth of the mouse neocortex, where the angioarchitecture has been previously described. First, we cross-validate average DyC-OCT cortical flow against conventional Doppler OCT flow. Next, with laminar DyC-OCT, we discover that layer 4 consistently exhibits the highest microvascular blood flow, approximately two-fold higher than the outer cortical layers. While flow differences between layers are well-explained by microvascular volume and density, flow differences between subjects are better explained by transit time. Finally, from layer-resolved tracer enhancement, we also infer that microvascular hematocrit increases in deep cortical layers, consistent with predictions of plasma skimming. Altogether, our results show that while the cortical blood supply derives mainly from the pial surface, laminar hemodynamics ensure that the energetic needs of individual cortical layers are met. The laminar trends reported here provide data that links predictions based on the cortical angioarchitecture to cerebrovascular physiology in vivo.

Laser capture microdissection-targeted mass spectrometry: a method for multiplexed protein quantification within individual layers of the cerebral cortex.

The mammalian neocortex is organized into layers distinguished by the size, packing density, and connectivity of their constituent neurons. Many neuropsychiatric illnesses are complex trait disorders with etiologic factors converging on neuronal protein networks. Cortical pathology of neuropsychiatric diseases, such as schizophrenia, is often restricted to, or more pronounced in, certain cortical layers, suggesting that genetic vulnerabilities manifest with laminar specificity. Thus, the ability to investigate cortical layer-specific protein levels in human postmortem brain is highly desirable. Here, we developed and validated a laser capture microdissection-mass spectrometry (LCM-MS) approach to quantify over 200 proteins in cortical layers 3 and 5 of two cohorts of human subjects as well as a monkey model of postmortem interval. LCM-MS was readily implementable and reliably identified protein patterns that differed between cortical layers 3 and 5. Our findings suggest that LCM-MS facilitates the precise quantification of proteins within individual cortical layers from human postmortem brain tissue, providing a powerful tool in the study of neuropsychiatric disease.

Two-photon microscopy measurement of cerebral metabolic rate of oxygen using periarteriolar oxygen concentration gradients

The cerebral metabolic rate of oxygen ([Formula: see text]) is an essential parameter for evaluating brain function and pathophysiology. However, the currently available approaches for quantifying [Formula: see text] rely on complex multimodal imaging and mathematical modeling. Here, we introduce a method that allows estimation of [Formula: see text] based on a single measurement modality-two-photon imaging of the partial pressure of oxygen ([Formula: see text]) in cortical tissue. We employed two-photon phosphorescence lifetime microscopy (2PLM) and the oxygen-sensitive nanoprobe PtP-C343 to map the tissue [Formula: see text] distribution around cortical penetrating arterioles. [Formula: see text] is subsequently estimated by fitting the changes of tissue [Formula: see text] around arterioles with the Krogh cylinder model of oxygen diffusion. We measured the baseline [Formula: see text] in anesthetized rats and modulated tissue [Formula: see text] levels by manipulating the depth of anesthesia. This method provides [Formula: see text] measurements localized within [Formula: see text] and it may provide oxygen consumption measurements in individual cortical layers or within confined cortical regions, such as in ischemic penumbra and the foci of functional activation.

Laminar analysis of the slow wave activity in the somatosensory cortex of anesthetized rats

Rhythmic slow waves characterize brain electrical activity during natural deep sleep and under anesthesia, reflecting the synchronous membrane potential fluctuations of neurons in the thalamocortical network. Strong evidence indicates that the neocortex plays an important role in the generation of slow wave activity (SWA), however, contributions of individual cortical layers to the SWA generation are still unclear. The anatomically correct laminar profiles of SWA were revealed under ketamine/xylazine anesthesia, with combined local field potential recordings, multiple-unit activity (MUA), current source density (CSD) and time-frequency analyses precisely co-registered with histology. The up-state related negative field potential wave showed the largest amplitude in layer IV, the CSD was largest in layers I and III, whereas MUA was maximal in layer V, suggesting spatially dissociated firing and synaptic/transmembrane processes in the rat somatosensory cortex. Up-state related firing could start in virtually any layers (III-VI) of the cortex, but were most frequently initiated in layer V. However, in a subset of experiments, layer IV was considerably active in initiating up-state related MUA even in the absence of somatosensory stimulation. Somatosensory stimulation further strengthened up-state initiation in layer IV. Our results confirm that cortical layer V firing may have a major contribution to the up-state generation of ketamine/xylazine-induced SWA, however, thalamic influence through the thalamorecipient layer IV can also play an initiating role, even in the absence of sensory stimulation.

Laminar microvascular transit time distribution in the mouse somatosensory cortex revealed by Dynamic Contrast Optical Coherence Tomography

The transit time distribution of blood through the cerebral microvasculature both constrains oxygen delivery and governs the kinetics of neuroimaging signals such as blood-oxygen-level-dependent functional Magnetic Resonance Imaging (BOLD fMRI). However, in spite of its importance, capillary transit time distribution has been challenging to quantify comprehensively and efficiently at the microscopic level. Here, we introduce a method, called Dynamic Contrast Optical Coherence Tomography (DyC-OCT), based on dynamic cross-sectional OCT imaging of an intravascular tracer as it passes through the field-of-view. Quantitative transit time metrics are derived from temporal analysis of the dynamic scattering signal, closely related to tracer concentration. Since DyC-OCT does not require calibration of the optical focus, quantitative accuracy is achieved even deep in highly scattering brain tissue where the focal spot degrades. After direct validation of DyC-OCT against dilution curves measured using a fluorescent plasma label in surface pial vessels, we used DyC-OCT to investigate the transit time distribution in microvasculature across the entire depth of the mouse somatosensory cortex. Laminar trends were identified, with earlier transit times and less heterogeneity in the middle cortical layers. The early transit times in the middle cortical layers may explain, at least in part, the early BOLD fMRI onset times observed in these layers. The layer-dependencies in heterogeneity may help explain how a single vascular supply manages to deliver oxygen to individual cortical layers with diverse metabolic needs.

Architecture and morphology of the human ventromedial prefrontal cortex.

A previous report identified the location of comparable architectonic areas in the ventral frontal cortex of the human and macaque brains [S. Mackey & M. Petrides (2010) Eur. J. Neurosci., 32, 1940-1950]. The present article provides greater detail with regard to the definition of architectonic areas within the ventromedial part of the human ventral frontal cortex and describes their location: (i) in Montreal Neurological Institute proportional stereotactic space; and (ii) in relation to sulcal landmarks. Structural magnetic resonance scans of four brains were obtained before the preparation of the histological specimens, so that the architectonic parcellation could be reconstructed in its original three-dimensional volume. The areal density of individual cortical layers was sampled quantitatively in the ventromedial prefrontal cortex of eight brains (16 hemispheres). The agranular cortex along the ventral edge of the corpus callosum and posterior margin of the ventromedial surface is replaced by a graded series of increasingly granular and more complexly laminated areas that succeed one another in a posterior-to-anterior direction. In parallel, the width of the supragranular layers (i.e. layers II and III) increases as compared with the infragranular layers (i.e. layers V and VI) from posterior to anterior. A measure of how rapidly cortical features change at areal boundaries also showed that the rate of change in the granule and pyramidal cell densities of layers IV and V, respectively, was greater at the borders between posterior areas than between anterior areas. This article will facilitate the anatomical identification and comparison of experimental data involving the human vmPFC.

Estrogen-related receptor α in normal adrenal cortex and adrenocortical tumors: Involvement in development and oncogenesis

AimsThe nuclear hormone receptor estrogen-related receptor α (ERRα) regulates the activation of mitochondrial genes in various human tissues, but its role in the adrenal gland and its disorders has not been defined. Therefore, we examined ERRα expression in both normal adrenal cortex (NAC) and adrenocortical tumor (ACT) in order to study the possible correlation of ERRα with adrenal development and tumor development. MethodsHuman adrenal specimens (non-pathological fetal n=7; non-pathological post-birth n=40; aldosterone producing adenoma (APA) n=11; cortisol producing adenoma (CPA) n=11; adrenocortical carcinoma (ACC) n=8) were immunohistochemically examined in this study. NAC (n=13) and ACT (n=28) frozen tissue specimens were also available for studying ERRα mRNA levels. Key findingsIn fetal NAC tissues, ERRα labeling index (LI) in fetal zone (FZ) was significantly higher that that in neocortex (NC), and the differences among age groups for overall mean LI was statistically significant when analyzed according to individual cortical layers. ERRα LI was also significantly higher in ACC than in other types of ACT. ERRα mRNA was detected in NAC and all types of ACT. SignificanceResults of our present study suggest a possible role of ERRα in adrenal development and ACC.

Transcriptional Architecture of the Primate Neocortex

Genome-wide transcriptional profiling was used to characterize the molecular underpinnings of neocortical organization in rhesus macaque, including cortical areal specialization and laminar cell-type diversity. Microarray analysis of individual cortical layers across sensorimotor and association cortices identified robust and specific molecular signatures for individual cortical layers and areas, prominently involving genes associated with specialized neuronal function. Overall, transcriptome-based relationships were related to spatial proximity, being strongest between neighboring cortical areas and between proximal layers. Primary visual cortex (V1) displayed the most distinctive gene expression compared to other cortical regions in rhesus and human, both in the specialized layer 4 as well as other layers. Laminar patterns were more similar between macaque and human compared to mouse, as was the unique V1 profile that was not observed in mouse. These data provide a unique resource detailing neocortical transcription patterns in a nonhuman primate with great similarity in gene expression to human.

Quantitative demonstration of comparable architectonic areas within the ventromedial and lateral orbital frontal cortex in the human and the macaque monkey brains

The orbital and ventromedial frontal cortical regions of the human and the macaque monkey brains include several spatially discrete areas which are defined histologically by their distinctive laminar architecture. Although considerable information has been collected on the function and anatomical connections of specific architectonic areas within the orbital and ventromedial frontal cortex of the macaque monkey, the location of comparable areas in the human brain remains controversial. We re-examined the comparability of orbital and ventromedial frontal areas across these two species and provide the first quantitative demonstration of architectonically comparable cortical areas in the human and the macaque brains. Images of Nissl-stained sections of the cortex were obtained at low magnification. Differences in the typical size of neurons in alternating pyramidal and granule cell layers were exploited to segregate the cortical layers before sampling. Profiles of areal neuronal density were sampled across the width of the cortex. The location of individual cortical layers was identified on each profile by sampling a set of equally sized images on which the cortical layers had been manually traced. The rank order of sampled architectonic features in comparable architectonic areas in the two species was significantly correlated. The differences in measured features between gyral and sulcal parts of the same architectonic area are at a minimum 3-4 times smaller than the differences between architectonic areas for the areas examined. Furthermore, the quantified architectonic features arrange areas within the orbital and ventromedial frontal cortex along two dimensions: an anterior-to-posterior and a medial-to-lateral dimension. On the basis of these findings, and in light of known anatomical connections in the macaque, this region of the human cortex appears to comprise at least two hierarchically structured networks of areas.

Architectonic mapping of the medial region of the human orbitofrontal cortex by density profiles

The study of architectonic differentiation in the cortex is advanced by the articulation of objective definitions based on clear features of the cortical architecture. We adopted areal density profiles as a means of sampling the cortex. On the profiles, we isolated and quantified the density of individual cortical layers. These features may serve as criteria in objective definitions in a way that builds on qualitative observations found in the classical literature. A preprocessing procedure was introduced to overcome artefacts in the density profiles caused by the partial overlap of neighboring neuronal layers and cortical folding. We applied this method to the medial half of the orbital frontal cortex in specimens drawn from 10 human postmortem brain hemispheres. The measurements successfully confirmed the existence of several qualitatively observed areas (architectonic areas 14c, 14r, 11m, 11 and 13). The selection of specific sampling parameters was justified on the basis of simultaneous measurements of the cortical morphology which demonstrate its influence on the appearance of the cortical layers. We also examined the robustness of the measuring procedure by analyzing the outcome of varying systematically the sampling parameters. We describe here a novel method of sampling the cortex for architectonic analysis and demonstrate its application on histological sections obtained from the medial half of the human orbitofrontal cortex.

Observer‐independent analysis of high‐resolution MR images of the human cerebral cortex: In vivo delineation of cortical areas

Using high-resolution MRI, it is now possible to examine the living human cortex down to a resolution of less than 300 mum. Thus, in vivo imaging is now approaching the resolution that has been successfully used in histological analysis of the cerebral cortex for many years, e.g., low-magnification light microscopy. This allows unprecedented views of cortical microstructure that reflect defined histological features, specifically, individual cortical layers. As in histological brain mapping, it is possible to use the changes in the cortical lamination patterns to define individual cortical areas. This allows in vivo neuroanatomical maps to be generated for individual subjects and precise correlation of the results from functional imaging studies in these subjects with their own microanatomical information. To this end, we adapted the well-established observer-independent cytoarchitectonic mapping techniques for defining cortical borders based on changes in cortical lamination for in vivo parcellation of high-resolution structural MR images.

Areal and laminar variations in the vascularity of the visual, auditory, and entorhinal cortices of the developing rat brain

Understanding of place-specific cortical cerebrovascular changes after insult and injury depends on the detailed knowledge of the areal and laminar variations in cortical vascularity. The present study examines comparatively the developmental changes of the total vascular density and the density of capillaries and medium- and large-sized vessels in the primary visual cortex (Oc1), the primary auditory cortex (Te1), and the lateral entorhinal cortex (EntL) of the developing rat brain. Vascular networks in the three cortical areas were marked after transcardial perfusion of India ink and quantified with an image analysis system. Parameters examined exhibited (i) peculiar developmental time course within individual cortical layers and (ii) area- and age-dependent variations. Angioarchitecture in Te1 layers was stabilized earlier than that in Oc1 layers and the period of postnatal development of the vascularity of neocortical sensory areas Oc1 and Te1 appeared to be more protracted compared to that of the phylogenetically older entorhinal cortex. By the end of the first postnatal month, vascular densities in the three cortical areas established a dorsoventral gradient (Oc1 > Te1 > EntL). Finally, in all areas, layer IV was the first layer to obtain adult values of capillary density.

Suberin lamella development in maize seedling roots grown in aerated and stagnant conditions

ABSTRACTHypoxia can stimulate the development of a suberized exodermis in aquatic plants; however, its influence on this aspect of terrestrial root development is sparsely documented. To determine the effects of hypoxia on maize (Zea mays cv. Seneca Horizon) roots, seedlings were grown in vermiculite (VERM), aerated hydroponics (AER), stagnant hydroponics with agar (STAG), or aerated hydroponics with agar (AERAG). The endo‐ and exodermis were examined for wall modifications. Lateral root emergence and aerenchyma formation were documented qualitatively. The endodermal Casparian band formation was unaffected by treatment. Endodermal and exodermal suberin lamella formation was earliest and most extensive in VERM. Suberization, especially in the exodermis of aerated treatments, was depressed in all hydroponic media. In comparison with AER, STAG exodermal lamellae were increased, but endodermal lamellae were decreased. Since the suberized exodermis forms a barrier to radial oxygen loss from roots to the medium, its stimulation in STAG roots (which also developed extensive aerenchyma) would help retain oxygen in the root. The reduction of endodermal lamellae should facilitate oxygen diffusion into the stele. Clearly, the response to environmental conditions is variable within individual cortical cell layers. Additionally, the observed patterns of lamellae, aerenchyma and lateral root development indicate a tight radial co‐ordination of root development.

Increased expression of insulin-like growth factor-I (IGF-I) during embryonic development produces neocortical overgrowth with differentially greater effects on specific cytoarchitectonic areas and cortical layers

The in vivo actions of insulin-like growth factor-I (IGF-I) on the growth and development of the cerebral cortex were investigated in transgenic (Tg) mice that overexpress IGF-I in the brain, beginning as early as embryonic day (E) 13. Compared to non-Tg littermate controls, Tg mice at postnatal day (P) 12 exhibited significant increases in total cortical volume (31%) and in total neuron number (27%). The numerical density of neurons did not differ significantly between Tg and control mice, except in layer I. Comparing cytoarchitectonic areas in Tg mice, significantly greater increases in cortical volume were found for the motor cortex (42%), compared to somatosensory cortex (35%). Similarly, greater increases in total neuron number were found for motor cortex (44%) compared to somatosensory cortex (28%). Comparing individual cortical layers in Tg mice, the greatest increase in neuron number was found in layer I for both motor (93%) and somatosensory (76%) regions, followed by layer V (36–53%)>II/III (26–47%)>VI (26–37%)>IV (22–34%). Our results demonstrate that increased expression of IGF-I in vivo during embryonic and early postnatal development produces substantial overgrowth of the neocortex. IGF-I-mediated growth and development exhibits differential effects in some cytoarchitectonic areas and in lamina-specific neuron populations, most notably the neurons of layer I.

Orientation Selectivity in Macaque V1: Diversity and Laminar Dependence

We studied the steady-state orientation selectivity of single neurons in macaque primary visual cortex (V1). To analyze the data, two measures of orientation tuning selectivity, circular variance and orientation bandwidth, were computed from the tuning curves. Circular variance is a global measure of the shape of the tuning curve, whereas orientation bandwidth is a local measure of the sharpness of the tuning curve around its peak. Circular variance in V1 was distributed broadly, indicating a great diversity of orientation selectivity. This diversity was also reflected in the individual cortical layers. However, there was a tendency for neurons with high circular variance, meaning low selectivity for orientation, to be concentrated in layers 4C, 3B, and 5. The relative variation of orientation bandwidth across the cortical layers was less than for circular variance, but it showed a similar laminar dependence. Neurons with large orientation bandwidth were found predominantly in layers 4C and 3B. There was a weak correlation between orientation selectivity and the level of spontaneous activity of the neurons. We also assigned a response modulation ratio for each cell, which is a measure of the linearity of spatial summation. Cells with low modulation ratios tended to have higher circular variance and bandwidth than those with high modulation ratios. These findings suggest a revision to the classical view that nonoriented receptive fields are principally found in layer 4C and the cytochrome oxidase-rich blobs in layer 2/3. Instead, a broad distribution of tuning selectivity is found in all cortical layers, and neurons that are weakly tuned for orientation are ubiquitous in V1 cortex.

Orientation selectivity in macaque V1: diversity and laminar dependence.

We studied the steady-state orientation selectivity of single neurons in macaque primary visual cortex (V1). To analyze the data, two measures of orientation tuning selectivity, circular variance and orientation bandwidth, were computed from the tuning curves. Circular variance is a global measure of the shape of the tuning curve, whereas orientation bandwidth is a local measure of the sharpness of the tuning curve around its peak. Circular variance in V1 was distributed broadly, indicating a great diversity of orientation selectivity. This diversity was also reflected in the individual cortical layers. However, there was a tendency for neurons with high circular variance, meaning low selectivity for orientation, to be concentrated in layers 4C, 3B, and 5. The relative variation of orientation bandwidth across the cortical layers was less than for circular variance, but it showed a similar laminar dependence. Neurons with large orientation bandwidth were found predominantly in layers 4C and 3B. There was a weak correlation between orientation selectivity and the level of spontaneous activity of the neurons. We also assigned a response modulation ratio for each cell, which is a measure of the linearity of spatial summation. Cells with low modulation ratios tended to have higher circular variance and bandwidth than those with high modulation ratios. These findings suggest a revision to the classical view that nonoriented receptive fields are principally found in layer 4C and the cytochrome oxidase-rich blobs in layer 2/3. Instead, a broad distribution of tuning selectivity is found in all cortical layers, and neurons that are weakly tuned for orientation are ubiquitous in V1 cortex.

Laminar distribution of nicotinic receptor subtypes in cortical regions in schizophrenia

The laminar cortical distribution of the [ 125I]α-bungarotoxin, [ 3H]cytisine and [ 3H]epibatidine nicotinic acetylcholine receptor ligands was investigated by quantitative autoradiography in autopsy tissue from the cingulate, orbitofrontal and temporal cortices of control and schizophrenia subjects matched for age and smoking history. Different laminar binding patterns were observed for the various nicotinic ligands both in schizophrenic and control brains. [ 125I]α-Bungarotoxin binding was distributed homogeneously across all cortical layers in all three brain regions, with highest binding densities in the cingulate cortex. [ 3H]Cytisine and [ 3H]epibatidine binding varied across the cortical ribbon, with high binding in layers I, III, V and VI, within the three cortical regions. A significantly reduced [ 125I] α-bungarotoxin binding (−54%) was observed in the cingulate cortex of schizophrenia subjects, in comparison with normal individuals who smoked tobacco. In the same brain region also a significantly higher [ 3H]cytisine binding (48–77%) was observed in nearly all layers, except for layer I of the schizophrenia subjects, when compared to normal individuals with a history of tobacco use. No significant changes in [ 3H]epibatidine binding was observed within the individual cortical layers between control subjects and patients with schizophrenia, but when calculated as a whole region (i.e. measurements performed across the whole cortical ribbon), the temporal cortex showed a significant increase in [ 3H]epibatidine binding in schizophrenia subjects compared to control subjects. The results suggest opposite changes of the α4β2 and α7 nicotinic receptor subtypes in the cingulate cortex of patients with schizophrenia which might reflect involvement of two different nicotinic receptor mechanisms in schizophrenia brain.

Identification of multiple somatostatin receptors in the rat somatosensory cortex during development.

The present study was aimed at identifying somatostatin receptor subtypes on the basis of their ligand-binding properties in the rat somatosensory cortex during fetal and postnatal development. Characterization of somatostatin-binding sites was performed in individual cortical layers by using three radioligands and eight competitors with known selectivities for the five somatostatin receptor subtypes. Binding sites sensitive to sst2-selective ligands were detected with high densities in the intermediate zone of the fetal cortex. From embryonic day 21 to 21 days postnatal (P21), mixed populations of receptors were detected in the cortical plate and emerging layers I-VI. Putative sst2 receptors were detected throughout the entire period but displayed different affinities for somatostatin and analogs, and a different sensitivity to GTP, depending on the developmental stage and the cortical layer considered. High densities of binding sites exhibiting characteristics of the sst1, sst3/5, and sst4 receptor subtypes were observed from P4 to P7, P7 to P14, and P7 to P21, respectively. In addition, each type of site exhibited a particular distribution pattern across the cortical layers that varied during the development. In the adult cortex, binding sites with sst1 and sst2 receptor characteristics were predominant. This study provides evidences of developmental expression windows of four sst receptor subtypes in selected areas of the rat cerebral cortex.